Alpha-naphthylisothiocyanate causes neutrophils to release factors that are cytotoxic to hepatocytes.
Administration of alpha-naphthylisothiocyanate (ANIT) to rats causes acute liver injury characterized in part by hepatocellular damage and marked neutrophil infiltration, effects that resemble drug-induced cholangiolitic hepatitis in people. ANIT-induced liver injury is neutrophil dependent. Moreover, ANIT can activate neutrophils in vitro. Since neutrophil-derived proteases can mediate hepatocellular killing, we hypothesized that ANIT stimulates neutrophils to release proteolytic enzymes that are cytotoxic to hepatic parenchymal cells. To test this hypothesis, neutrophils were isolated from Sprague-Dawley rats and incubated with ANIT for 6-24 h. ANIT (6-50 microM) was not toxic to neutrophils as indicated by the lack of lactate dehydrogenase release into the incubation medium. The conditioned medium from ANIT-treated neutrophils (ANCM) was collected, centrifuged, added to isolated hepatocytes, and incubated for 8, 16, or 24 h. Conditioned medium collected from neutrophils exposed to 25 or 50 microM ANIT for 16-24 h caused hepatocellular damage as indicated by the release of alanine aminotransferase into the culture medium. The concentration of ANIT in ANCM was nondetectable (0.5 microM). Analysis of ANCM indicated the presence of both cathepsin G and elastase activities. Inhibitors of these enzymes afforded protection against ANCM-induced hepatocellular injury. These results indicate that ANIT causes neutrophils to release toxic proteases which cause hepatocellular damage in vitro.
Hill
D A
DA
Department of Pharmacology, Michigan State University, East Lansing 48824, USA.
Roth
R A
RA
D015058:1-Naphthylisothiocyanate / Q000633:toxicity*; D000410:Alanine Transaminase / Q000378:metabolism; D000818:Animals; D056649:Cathepsin G; D002403:Cathepsins / Q000378:metabolism; D002470:Cell Survival; D002478:Cells, Cultured / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D017077:Culture Media, Conditioned; D019272:Leukocyte Elastase / Q000378:metabolism; D008099:Liver / Q000378:metabolism / Q000473:pathology*; D008297:Male; D009504:Neutrophils / Q000187:drug effects* / Q000201:enzymology; D051381:Rats; D017207:Rats, Sprague-Dawley; D012697:Serine Endopeptidases / Q000378:metabolism*
D017077:Culture Media, Conditioned; D015058:1-Naphthylisothiocyanate; D000410:Alanine Transaminase; D002403:Cathepsins; D012697:Serine Endopeptidases; D056649:Cathepsin G; C534097:Ctsg protein, rat; D019272:Leukocyte Elastase